Abstract Sex differences are evident in vulnerability to age-related cognitive decline and diseases of aging. Estradiol (E2) is protective against neurodegenerative diseases, including Alzheimer?s disease, implicating sex hormone effects on sex differences in vulnerability. However, obstacles to sex steroid treatments include closing of the therapeutic window observed as decreased effectiveness of E2 treatment with advanced age. The goal of the proposed research is to provide an understanding of the mechanisms for E2 effects on memory and the closing of the therapeutic window. Closing of the therapeutic window is marked by a decrease in E2-responseive transcription and an inability of E2 treatment to enhance N-methyl-D-aspartate receptor (NMDAR)-mediated synaptic transmission examined several days after treatment. Aim 1 will test the hypothesis that E2 treatment, several days prior to testing, specifically influences NMDAR-dependent episodic memory, such that it can rescue an age-related decline in episodic memory examined on the water maze and novel object recognition tasks. Aim 2 will test the hypothesis that E2 effects on memory and NMDAR function are mediated by reversal of NMDAR hypofunction, mediated by redox regulation of phosphatase/kinase activity, similar to that previously described in aging males. Thus, it is predicted that prior to closing of the therapeutic window (i.e. in animals in which E2 treatment improves cognition and increases NMDAR function), E2 treatment will promote antioxidant enzyme activity, reduce oxidative stress, and minimize redox-mediated decrease in CaMKII activity and NMDAR function. Further, following closing of the therapeutic window (i.e. for animals in which E2 does not rescue cognition and NMDAR function), E2 treatment will not promote antioxidant enzyme activity or reduce oxidative stress, and the NMDAR response and CaMKII activity will be decreased due to an oxidized redox state. Aim 3 will test the hypothesis that age- related changes in transcriptional responsiveness to E2 are due, at least in part, to epigenetic regulation through DNA methylation. It is predicted that decreased responsiveness of E2-sensitive genes will be associated with DNA hypermethylation, particularly in gene body regions (introns), and specific to CpG, relative to non-CpG methylation sites. The proposed studies will employ a powerful combination of behavioral tests that are sensitive to NMDAR function, patch-clamp recording of NMDAR synaptic responses, measures of oxidative stress and enzyme activity, transcription, and DNA methylation.